This ongoing project involves several studies. Efforts to increase the sensitivity of the shell vial assay in detecting cytomegalovirus (CMV) continue. We are in the process of examining the effect of thymidine analogues, such as indo- or bromo-deoxyuridine, on CMV growth to determine if they can be added to our shell vial cultures to enhance the sensitivity of the culture to detect CMV in clinical specimens. Hexadimethrine bromide, or polybrene, has been demonstrated to help human herpes virus 6 penetrate cells in culture and therefore increase the sensitivity of the assay. We will examine the effect of polybrene on CMV in shell vial cultures to see if it can increase the sensitivity of the shell vial assay. The detection of pp65, the early structural protein of CMV, has been found to be a rapid and sensitive means of predicting CMV disease progression. We are examining and evaluating a pool of monoclonal antibodies to pp65 that recognizes the epitopes expressed in the nuclei of infected peripheral polymorphonuclear leukocytes and monocytes and that are different from our currently used antibodies to pp65. It is hoped that these new antibodies will allow an increased detection sensitivity of this CMV antigen in clinical blood specimens. CMV can be a cause of pneumonitis in the patient with HIV. Detection of CMV in smears consisting of cells found in bronchoalveolar lavages can be a rapid means of diagnosing CMV pulmonary disease. We will examine such smears using immunofluorescent reagents for the detection of CMV pp65 and the CMV 72kd immediate early antigen to determine their clinical utility as rapid diagnostic methods.